Horizontal gene transfer (HGT) is a major driver of bacterial evolution, but its role in shaping the human gut microbiome over time remains poorly understood. Here, we present a longitudinal metagenomic analysis of 676 fecal samples from 338 individuals in the Lifelines-DEEP study collected ~4 years apart, using a newly developed workflow to detect recent HGT events from metagenome-assembled genomes. We identified 5,644 high-confidence HGT events occurring within the past ~10,000 years across 116 gut bacterial species. We find that species pairs with an HGT relationship were significantly more likely to maintain stable co-abundance relationships over the 4-year period, suggesting that gene exchange contributes to community stability. Notably, HGT and strain replacement act together to disseminate mobile genes in the population. Furthermore, our observation that an individual's mobile gene pool remains highly personalized and stable over time indicates that host lifestyles drive specific gene transfer. For example, proton pump inhibitor usage is linked to increased transfer of multidrug transporter genes. Our findings demonstrate, at the individual gut microbiome level, that HGT is both an integral and stabilizing force in the human gut ecosystem and an important mechanism for disseminating adaptive functions, underscoring HGT potential for tracking host lifestyle.

ObjectiveThe latest European Society of Gastroenterology Hepatology and Nutrition (ESPGHAN) criteria for celiac disease (CD) diagnosis reduced the requirement for a small intestinal biopsy but still, for most of the cases a small intestinal biopsy is required for a safe diagnosis: hence the attempt to identify serum biomarkers that could replace, in most of these latter cases, the requirement of the biopsy (what is called a "liquid biopsy"). The aim of this study is to identify a set of serum biomarkers able to differentiate celiac patients from age and human leukocyte antigen (HLA)-matched healthy controls.MethodsRelative concentration of 92 inflammation-linked proteins was examined in the sera of 50 children with CD compared with 50 HLA DQ2/8 and age-matched healthy controls born in genetically at-risk families, using proximity extension immunoassay technology (Olink Proteomics®) with the ProSeek Multiplex Inflammation panel.ResultsThree different multivariate analysis (Univariate and multivariate distribution analysis, random forest classification and linear discriminant analysis) localized a cluster of seven molecules (CASP8, CXCL9, NT-3, SIRT2, STAMBP, ST1A1, and TNFSF14) with a remarkable diagnostic potential, able to differentiate around 90% (95% confidence interval [CI]; 0.7-0.99) of CD patients from controls.ConclusionPatients with CD, compared to age- and HLA-matched healthy controls, show in their sera an increased expression of inflammatory molecules, involved in NF-κB cytokine signaling, cell apoptosis, and crypt proliferation pathways. A set of seven of these proteins can differentiate cases from controls with an accuracy higher than 90%. The implementation of this approach in clinical setting could in future facilitate a noninvasive and individualized approach for CD diagnosis.

Summary The human hippocampus is a critical brain region for learning, memory, and stress regulation, distinguished by its ability to sustain neurogenesis after birth. This plasticity is driven by hippocampal neural stem cells (NSCs), which generate new neurons and maintain circuit integrity, but are highly sensitive to environmental and pathological influences. Mechanistic insight into human hippocampal development and neurogenesis remains limited by the absence of physiologically relevant models. Here, we establish an optimized protocol to generate human induced pluripotent stem cell-derived hippocampal organoids that recapitulate key features of hippocampal development. These organoids maintain organized NSC niches, support ongoing neurogenesis, and generate hippocampus-specific cell types. Cellular, transcriptomic, and electrophysiological analyses confirm progressive neuronal maturation, synapse formation, and functional activity, highlighting the physiological relevance of the system. Using this model, we modeled excess prenatal glucocorticoid exposure with dexamethasone, which perturbed NSC dynamics by reducing proliferation and inducing a precocious quiescent-like state. RNA sequencing revealed downregulation of NSC activation genes and upregulation of quiescence- and autophagy-associated programs, suggesting that glucocorticoid signaling enforces an early transition toward quiescence. These findings reveal a mechanism by which excessive glucocorticoid exposure may impair hippocampal growth. Together, this study introduces a robust human hippocampal organoid platform for dissecting the regulation of hippocampal development and for modeling the impact of environmental stressors on human neurogenesis.

BackgroundHuman milk dynamically adapts its composition of immunoglobulins (Igs), cytokines, and other proteins as lactation progresses, influencing the infant's immune development and protection. Understanding how maternal factors, such as parity, influence the composition of human milk can provide strategies aimed at enhancing infant immune protection and reducing early-life infections. This study aims to investigate whether the immune composition of human milk differs based on parity, and if so, how these changes are related to infections in early life.MethodsThe study included 75 healthy mother-infant pairs from the MAMI cohort (Clinical Trial Registry NCT03552939), with milk samples collected from the same mothers at days 7 and 15 postpartum, during transitional lactation stage. Igs, cytokines, and adipokines were quantified using multiplex immunoassays and ELISA. A comparison was conducted between primiparous and multiparous mothers regarding both the overall and individual composition of immune components in human milk at each time point, as well as their evolution throughout the transitional phase.ResultsInfants from multiparous mothers recorded higher infection rates in early life than those of primiparous mothers. Some human milk immune components also differed by parity, with multiparous mothers exhibiting higher levels of IgA, total IgG, IgG1, IgG2, IgG3, IgE, and IL-23 at the beginning of the transitional phase (day 7), as well as higher IL-18 and IL-21 levels toward its end (day 15), compared to primiparous mothers. Additionally, the evolutionary pattern in levels of Igs, cytokines, and adipokines throughout the transitional milk stage also differed. Moreover, in multiparous mothers, higher levels of IgG, particularly IgG1 and IgG2 (day 7), as well as IL-18 and IL-22 (day 15), were associated with reduced infant infections, highlighting their potential protective role.ConclusionsParity is a maternal factor that influences some immune components of human milk during the transitional stage and may be linked to the susceptibility of infants to infections during the first 6 months of life. Future studies aimed at analyzing the impact of the parity factor, among others, on the progression of immune components in human milk may contribute to a better understanding and improved strategies for newborn health.

Diagnosing celiac disease (CD) via esophagogastroduodenoscopy (EGDS) is necessary when anti-transglutaminase (anti-TG) antibody levels are below 10× the upper limit of normal (ULN). This study evaluates patients with low anti-TG titers, particularly when endomysial antibodies (EMA) are negative. In this retrospective study (2022-2024), patients undergoing EGDS for suspected CD were grouped by EMA status: Group 1 (EMA negative) and Group 2 (EMA positive), with similar low anti-TG titers. Group 1 (N = 25) had a mean anti-TG titer of 1.86× ULN and villous atrophy (VA) in only 8% (2/25). Group 2 (N = 100) had VA in 35% (35/100), a 6.16-fold higher risk. Nonatrophic cases showed no significant histological and immunohistochemical differences. In conclusion, low-titer anti-TG with negative EMA indicates a low likelihood of VA. Most asymptomatic patients may not require immediate intervention but should be monitored. EGDS can be reserved to later stages if clinical suspicion persists.

Early-life gut microbiome-metabolome crosstalk plays a crucial role in maintaining host physiology. The microbially produced metabolites often convey effects on host health and physiology. This study investigates the gut metabolites, including short-chain fatty acids (SCFAs), bile acids (BAs), and polar metabolites, and their relationship to gut microbiota composition in a birth cohort of 670 children. Samples were collected at 2.5 (n = 272), 6 (n = 232), 14 (n = 289), and 30 months (n = 157) of age. We identified the trajectories of the fecal metabolome that relate to the maturation of the early-life gut microbiota. We found that prevalent gut microbial abundances were associated with microbial metabolite levels, particularly in 2.5-month-old infants. Here, the abundances of early colonizers, e.g., Bacteroides, Escherichia, and Bifidobacterium, were associated with microbial metabolites, especially secondary BAs, particularly in breastfed infants. Our results suggest that early-life gut microbiota associates with changes in metabolome composition, particularly BAs, which may have physiological implications.

Human development and physiology are fundamentally linked with the microbiome. This is particularly true during early life, a critical period for microbiome assembly and its impact on the host. Understanding microbial acquisition in early life is thus central to both our basic understanding of the human microbiome and strategies for disease prevention and treatment. Here, we review the historical approaches to categorize microbial transmission originating from the fields of infectious disease epidemiology and evolutionary biology and discuss how this lexicon has influenced our approach to studying the early-life microbiome, often leading to confusion and misinterpretation. We then present a conceptual framework to capture the multifaceted nature of human microbiome acquisition based on four key components: what, where, who, and when. We present ways these parameters may be assigned, with a particular focus on the 'transmitted strain' through metagenomics to capture these elements. We end with a discussion of approaches for implementing this framework toward defining each component of microbiome acquisition.

Human milk is important for infant development, but few large studies have comprehensively investigated milk composition. Here, we characterized human milk oligosaccharides (HMOs) and milk microbiota, their shaping factors, and their links to infant gut microbiota in the longitudinal Dutch Lifelines NEXT cohort. We measured 24 HMOs in 1,542 milk samples from 524 mothers at 0.5-6 months postpartum, profiled microbiota in milk and maternal and infant feces, genotyped mothers, and recorded 174 environmental, maternal, and infant characteristics. HMO concentrations were associated with maternal genetic loci (FUT2, FUT3/FUT6, ABO, and ST3GAL6), lactation stage, and subclinical mastitis. The human milk microbiota varied during lactation and with different feeding practices. Both HMOs and milk microbiota remained stable across multiple pregnancies in the same individual. Some milk bacteria were present in infant feces, but the milk and infant fecal microbiota diverged as the infant aged. Furthermore, individual HMOs were associated with infant fecal microbiota characteristics.

BackgroundMaternal nutritional status and dietary profile during pregnancy and lactation have short- and long-term impacts on offspring health. However, there is an incomplete understanding of the mechanisms behind these health effects. This study aims to assess the effect of maternal diet on the health of offspring by examining to unravel the impact of maternal diet on offspring health outcomes and evaluate the link between maternal nutrition, human milk immune components and neonatal colonisation as potential mechanisms that mediate the influence of maternal diet in the incidence of infant infections.MethodsTo assess this objective, we used two complementary approaches by which a clinical observational study based on the MAMI birth cohort guided a preclinical interventional analysis using a neonatal rat model of rotavirus-induced gastroenteritis.FindingsThe findings in both approaches demonstrated that a maternal diet rich in plant-based protein, fibre and polyunsaturated fatty acids, was linked to reduced incidence and severity of infections in offspring that would be mediated by beneficial modulation of the gut microbiota and immune system. Specifically, in the suckling rats, a predominant Th1 immune response and an enhanced virus-specific response were observed. Moreover, human milk IgA and rat milk IgG2c played a key protective role that complemented the effects of maternal diet.InterpretationThese results strengthen the importance of maternal diet during pregnancy and lactation supporting infant health.FundingThe study was supported by LaMarató-TV3 (DIM-2-ELI, ref. 2018-27/30-31).

Contaminant sequences of external origin complicate the study of host-associated viromes, particularly in low-biomass samples obtained through viral-like particle (VLP) enrichment. However, the prevalence and impact of external contaminants on low-biomass samples are under-studied. Here, we analyze 1321 gut virome samples and 55 negative controls (NCs) from four early-life virome studies. Virus sequences identified in NCs, termed negativeome, were used as a proxy for the contamination assessment. We show that 61% of samples share at least one identical strain with negativeome, likely representing external contamination. While the median abundance of contaminant strains in these samples is only 1%, it ranges from 0 to 99% and exceeds 10% in 11% of infant samples. We further demonstrate that contamination is largely study-specific and has a greater impact on infant samples than on maternal samples. Based on our results, we propose a contamination assessment method using a publicly available database of sequences detected in NCs and a strain-level decontamination strategy.